N-nitrosamines have become of more active concern in recent times due to the recent technology revealing that some of these compounds are active carcinogens. These compounds have been found in many common foods. For example, N-nitrosamines have been found in nitrite treated meats such as bacon, and recent reports have discovered their presence in beverages such as scotch and beer. There is, therefore, a continuing need for techniques to determine, both quantitatively and qualitatively, the N-nitrosamine levels in a variety of compositions to which man is continually exposed.
N-nitrosamines have the following basic structures: ##STR1## wherein both R and R.sub.1 may represent a variety of organic or substituted organic moieties. In analyzing such compounds to determine not only their presence but also the level of concentration of the compounds present in any test sample, the bond between the nitrogen and the NO moiety is cleaved to produce nitric oxide gas which is removed from the reaction solution.
This bond is cleaved in an acid catalyzed reaction. Commonly in the past, this denitrosation reaction has employed as the reagents, glacial acetic acid with HBR; see, for example, Eisenbrand, G., et al., Arzneim-Forsch, 20, 1513 (1970). This is, however, a slow reaction and the presence of water in the test sample interferes with the reaction to substantially prevent liberation of nitric oxide gas. Other reported methods include those of Drescher, S., and Frank, C. W. Anal. Chem., 50, 2118-2121 (1978) and Downes, M. J. Edwards, M. W., Elseg, T. S., and Walters, C. L., Analyst 101 742 (1976). The Drescher et al. method employs methylene chloride solvent, glacial acetic acid and HBR. The Downes, et al. method employs 1,2 dichloroethane solvent.
In these methods, as well as other methods of denitrosation apparently being used, a significant problem is that the reactions employed are intolerant to even small amounts of water present in the reaction system or the test samples. That is to say, the presence of water inhibits the cleavage reaction from occurring and accordingly, insufficient amounts of nitric oxide are released within a reasonable period of time to conduct the analysis.
Accordingly, a primary object of this invention is to develop a denitrosating reagent composition which is usable even in the presence of water to effectively cleave the N--NO bond of an N-nitrosamine.
Another object of this invention is to provide an improved denitrosating reagent which not only is usable in the presence of water, but also reacts quickly to provide NO gas for further use in the analysis being conducted.
An even further object of this invention is to provide a method of denitrosating N-nitrosamines which employs the improved denitrosating reagent compositions of the invention.
Another object of this invention is to provide an improved method of determination of N-nitrosamines in a test sample, which method is usable even in the presence of large amounts of water in the test sample.
A still further object of this invention is to provide an improved degassing cell which is specifically designed for use with small, microliter amounts of liquid, to efficiently and effectively strip gaseous components dissolved in the small amount of liquid.
An even further object of the invention is to provide a detection method, means, and apparatus for specific use with N-nitrosamine analysis which is quick to operate, efficient to operate, tolerant to water in the test samples, and which accurately allows qualitative and quantitative detection of N-nitrosamines.
The method composition means and device for accomplishing the stated objectives of this invention, as well as others, will become apparent from the detailed description of the invention which follows.